%0 Journal Article %A Bansal, Ankita %A Kar, Debasish %A Murugan, Rajagopal A. %A Mallick, Sathi %A Dutta, Mouparna %A Pandey, Satya Deo %A Chowdhury, Chiranjit %A Ghosh, Anindya S. %T A putative low-molecular-mass penicillin-binding protein (PBP) of Mycobacterium smegmatis exhibits prominent physiological characteristics of dd-carboxypeptidase and beta-lactamase %D 2015 %J Microbiology, %V 161 %N 5 %P 1081-1091 %@ 1465-2080 %R https://doi.org/10.1099/mic.0.000074 %I Microbiology Society, %X dd-Carboxypeptidases (dd-CPases) are low-molecular-mass (LMM) penicillin-binding proteins (PBPs) that are mainly involved in peptidoglycan remodelling, but little is known about the dd-CPases of mycobacteria. In this study, a putative dd-CPase of Mycobacterium smegmatis, MSMEG_2433 is characterized. The gene for the membrane-bound form of MSMEG_2433 was cloned and expressed in Escherichia coli in its active form, as revealed by its ability to bind to the Bocillin-FL (fluorescent penicillin). Interestingly, in vivo expression of MSMEG_2433 could restore the cell shape oddities of the septuple PBP mutant of E. coli, which was a prominent physiological characteristic of dd-CPases. Moreover, expression of MSMEG_2433 in trans elevated beta-lactam resistance in PBP deletion mutants (ΔdacAdacC) of E. coli, strengthening its physiology as a dd-CPase. To confirm the biochemical reason behind such physiological behaviours, a soluble form of MSMEG_2433 (sMSMEG_2433) was created, expressed and purified. In agreement with the observed physiological phenomena, sMSMEG_2433 exhibited dd-CPase activity against artificial and peptidoglycan-mimetic dd-CPase substrates. To our surprise, enzymic analyses of MSMEG_2433 revealed efficient deacylation for beta-lactam substrates at physiological pH, which is a unique characteristic of beta-lactamases. In addition to the MSMEG_2433 active site that favours dd-CPase activity, in silico analyses also predicted the presence of an omega-loop-like region in MSMEG_2433, which is an important determinant of its beta-lactamase activity. Based on the in vitro, in vivo and in silico studies, we conclude that MSMEG_2433 is a dual enzyme, possessing both dd-CPase and beta-lactamase activities. %U https://www.microbiologyresearch.org/content/journal/micro/10.1099/mic.0.000074